The ultimate goal of oncologic cellular therapy is to infuse patients with a high number of potent tumor- reactive T lymphocytes. Both the quantity and quality of antitumor T cells (usually CD8+ T cells) transferred into a cancer patient are integral to long-term tumor remission. Unfortunately, therapeutic success is limited by the phenomenon that tumor-reactive CD8+ T cells lose their antitumor capacity as they are expanded to large populations in vitro. Until recently, all T cells were thought to experience a similar decline in antitumor potency as they matured through ex vivo expansion. Yet, because CD4+ T cells that secrete IL-17A, called Th17 cells, are capable of self-renewal using stem-cell associated pathways, it is possible that Th17 cells may not play by these rules. To test this hypothesis, Th17 cells were expanded to days 6, 12, and 18 before adoptive transfer into mice with large, established melanoma tumors. Not only did Th17 cells expanded to greater than 10,000 fold by day 18, but day 18 Th17 cells were shockingly as effective at eradicating melanomas as day 6 and 12 Th17 cells. We propose 2 aims to define and translate the mechanism responsible for the robust expansion and sustained antitumor potency of Th17 cells. First, we will test the hypothesis that the intrinsic mechanism that sustains the Th17 antitumor response is mediated by high expression of an anti-apoptotic molecule called c- FLIP. We posit that loss of this molecule will result in abrogation of the preserved antitumor efficacy of Th17 cells. Second, we seek to determine if human Th17 cells also maintain their capacity to elicit potent antitumor responses after 18 days of expansion. We will also investigate a second strategy that c-FLIP overexpression in human primary blood lymphocytes (PBLs) will endow them with longevity over ex vivo expansion and preserve their antitumor response. The mouse experiments in Aim 1 will be performed using CD4+ T cells from transgenic TRP-1 mice, and CD8+ T cells from transgenic Pmel-1 mice. Aim 2 will use humanized models using chimeric antigen receptor (CAR) transduced human T cells against human tumors in NSG immunocompromised mice. As maintaining the therapeutic quality of T cells that are expanded to large clinically relevant numbers is a main goal of cancer immunotherapy field, this proposed research will reveal significant insights into the role and application of Th17 cells in tumor immunity.